Esophageal cancer is the eighth most prevalent cancer, and the sixth leading cause of cancer death worldwide. There are two histologic types of esophageal cancer; squamous cell carcinoma (SCC) and adenocarcinoma. SCC is by far the most common type worldwide, accounting for about 90% of all esophageal cancer. This disease has a poor prognosis with a 5-year survival rate of about 12%. The major risk factors for esophageal SCC are tobacco and alcohol use, consumption of salty and moldy foods that are frequently contaminated with N-nitrosamine carcinogens, vitamin and trace mineral deficiencies, and diets high in starch but low in fruits and vegetables. Our laboratory has been evaluating the use of freeze-dried berries, mainly black raspberries (BRBs), for the prevention of esophageal squamous cell carcinoma in rats. In the rat, esophageal tumors are induced within 25-30 weeks by repeated s.c. injections of the N-nitrosamine carcinogen, N-nitrosomethylbenzylamine (NMBA). The addition of 5% and 10% BRB powder to the diet of NMBA-treated rats reduces the number of esophageal tumors by 50-60%. During the past 5 years, we demonstrated that BRBs reduce NMBA-induced DNA damage in the rat esophagus and they inhibit proliferation, inflammation and angiogenesis, and stimulate apoptosis in premalignant esophageal cells and tissues. They also modulate the expression levels of genes associated with all of these cellular functions. Using bio-directed fractionation, we and our collaborators found that the anthocyanins in BRBs are important for their chemopreventive effects. The anthocyanins are responsible for the color of BRBs and much of their antioxidant activity. Because we rarely observe more than 50-60% inhibition of esophageal carcinogenesis with BRBs or the anthocyanins alone, we now propose to use the anthocyanins in combination with phenethyl isothiocyanate (PEITC) for the prevention of esophageal cancer. In the 1990's we identified PEITC as a very potent inhibitor of tumor initiation in the rat esophagus, however, unlike BRBs, PEITC failed to inhibit esophageal tumor progression when given in the diet post-initiation. We propose therefore, to identify low dietary concentrations of BRB anthocyanins (Specific Aim 1) and of PEITC (Specific Aim 2) that inhibit NMBA- induced esophageal tumorigenesis, and then evaluate these agents in combination (Specific Aim 3) with the objective of achieving a high level of chemopreventive efficacy using minimal dietary levels of these agents. We also propose to examine the effects of dietary anthocyanins and PEITC, alone and in combination, on the formation of NMBA-induced DNA adducts in rat esophagus as well as their effects on the expression of genes associated with cell proliferation, inflammation, angiogenesis and apoptosis (Specific Aim 4). Ultimately, we plan to evaluate these agents for prevention of esophageal SCC in humans at high risk for the disease. PUBLIC HEALTH RELEVANCE: Esophageal cancer is the eighth most prevalent cancer, and the sixth leading cause of cancer death worldwide. Our laboratory is developing strategies for the prevention of esophageal cancer using freeze-dried berries and isothiocyanate compounds found in vegetables. We now propose to evaluate the anthocyanins in berries in combination with phenethyl isothiocyanate in vegetables for their ability to prevent esophageal cancer at low, non-toxic, dietary concentrations.